One thousand patients with essential thrombocythemia: the Mayo Clinic experience

We describe 1000 patients with essential thrombocythemia seen at the Mayo Clinic between 1967 and 2023: median age 58 years (18–90), females 63%, JAK2/CALR/MPL-mutated 62%/27%/3%, triple-negative (TN) 8%, extreme thrombocytosis (ExT; platelets ≥1000 × 109/L) 26%, leukocytosis (leukocyte count >11 × 109/L) 20%, and abnormal karyotype 6%. JAK2-mutated patients were older (median 71 years), and CALR mutated (52 years), and TN (50 years) younger (p < 0.01). Female gender clustered with TN (73%) and JAK2 (69%) vs. CALR/MPL (49%/47%) mutations (p < 0.01). ExT clustered with CALR (type-2 more than type-1) and TN and leukocytosis with JAK2 mutation (p < 0.01). In multivariable analysis, risk factors for overall survival were older age (p < 0.01), male gender (HR 1.8), absolute neutrophil count (ANC) ≥ 8 × 109/L (HR 1.6), absolute lymphocyte count (ALC) < 1.7 × 109/L (HR 1.5), hypertension (HR 1.7), and arterial thrombosis history (HR 1.7); for leukemia-free survival, ExT (HR 2.3) and abnormal karyotype (HR 3.1); for myelofibrosis-free survival, ANC ≥ 8 × 109/L (HR 2.3) and MPL mutation (HR 3.9); for arterial thrombosis-free survival, age ≥60 years (HR 1.9), male gender (HR 1.6), arterial thrombosis history (HR 1.7), hypertension (HR 1.7), and JAK2 mutation (HR 1.8); for venous thrombosis-free survival, male gender (HR 1.8) and venous thrombosis history (HR 3.0). Associations between ExT and leukemic transformation and between ANC and fibrotic progression were limited to JAK2-mutated cases. Aspirin therapy appeared to mitigate both arterial (HR 0.4) and venous (HR 0.4) thrombosis risk. HR-based risk models delineated patients with median survivals ranging from 10 years to not reached and 20-year leukemia/myelofibrosis incidences from 3%/21% to 12.8%/49%. The current study provides both novel and confirmatory observations of essential thrombocythemia.


INTRODUCTION
Essential thrombocythemia (ET) is one of four JAK2 mutationprevalent myeloproliferative neoplasms (MPNs) and is characterized by a mandatory but not specific thrombocytosis (platelet count ≥450 × 109/L) that is proven or presumed to be clonal and not associated with another myeloid neoplasm, such as chronic myeloid leukemia (CML), polycythemia vera (PV), and primary myelofibrosis (PMF) [1,2].The latter two share common molecular and morphologic traits with ET, including JAK2, CALR, and MPL mutations (also known as MPN driver mutations); these mutations are mutually exclusive, for the most part, and their frequencies in ET are ~60% for JAK2, 25% for CALR, and 3% for MPL; of note, these three driver mutations might not be detected in ~10-15% of patients with ET, henceforth referred to as triple-negative ET [1,3].
Prognosis in ET is generally favorable with consistent risk of thrombohemorrhagic complications and disease progression into myelofibrosis (post-ET MF) or acute myeloid leukemia (AML), also known as "blast phase MPN" [4][5][6].Survival in ET approximates that of the general population with median estimated to exceed 30 years in patients younger than 40 years of age [7][8][9].The recently introduced "triple A (AAA)" survival model in ET employs age, absolute neutrophil count, and absolute lymphocyte count, in order to risk-stratify patients into high, intermediate-2, intermediate-1, and low-risk groups, with respective median survivals of 8, 13.5, 20.7, and 47 years [10].In addition, abnormal karyotype [11] and high molecular risk (HMR-ET; SF3B1, TP53) mutations [12] independently predict inferior survival in ET.Current drug therapy has not been shown to modify the natural history of the disease, and its use is primarily directed at the prevention of thrombosis, guided by thrombosis risk models that are based on thrombosis history, age, and presence of JAK2 mutation [13,14].
Over the last half-century, the Mayo Clinic has been a center of excellence for patient care and research in MPN, under the leadership of the late Murray N. Silverstein ; part of this decades-long experience has been assembled into previously published large natural history studies, including a 1000-patient report on primary myelofibrosis [15].The current report includes 1000 patients with ET, seen at the Mayo Clinic between 1967 and 2023, and selected on the basis of full annotation for driver mutations; we describe presenting clinical and laboratory characteristics, frequency and outcome of post-diagnosis events, and detailed global and driver mutation-specified analyses of overall, leukemia-free, and myelofibrosis-free survival, as well as predictors for such events.

METHODS
The current study includes 1000 consecutive patients with ET, who underwent evaluation at the Mayo Clinic between December 1967 and March 2023 and in whom bone marrow biopsies and driver mutation information was available for review.All cases fulfilled the ICC 2022 diagnostic criteria [16] and were fully annotated for driver mutations, while cytogenetic information was available in a subset of patients (n = 875).Patients were retrospectively recruited after institutional review board approval was obtained.In order to minimize the inadvertent inclusion of patients with masked PV [17], JAK2 mutated cases with hemoglobin level >16 g/dL in women and 16.5 g/dL in men were excluded; similarly, cases with anemia defined by sex adjusted hemoglobin level of <11 g/dL in women and <12.5 g/dL in men without an alternative explanation were also excluded, in order to avoid inadvertent inclusion of patients with prefibrotic MF [18].Thrombosis and survival risk was assessed by the revised IPSET-thrombosis [13] and triple A survival model [10], respectively.Conventional criteria were used for definitions of major arterial and venous thrombotic events, major hemorrhage, fibrotic or leukemic transformation [4,6,16].Therapeutic interventions were dependent on physician discretion and mostly included aspirin therapy in low-risk patients and the addition of cytoreductive therapy, in high-risk patients.Patients were followed until death or last follow-up, as assessed by medical records or through direct contact with patients or their physicians, with follow-up information updated in August 2023.
Comparison between categorical variables was performed by Chi-square test and continuous variables by Wilcoxon/Kruskal-Wallis tests.Cox regression analysis was used to identify risk factors for overall (OS), leukemia-free survival (LFS), myelofibrosis-free (MFFS), and thrombosis-free (TFS).The Kaplan-Meier method was used to construct time-to-event curves, which were compared by the log-rank test.P value ≤ 0.05 was considered significant.JMP Pro 17.1.0software package, SAS Institute, Cary, NC was utilized for all analyses.
Aspirin therapy appeared to mitigate both arterial and venous thrombosis with arterial and venous thrombosis rates of 5% vs 16% and 3% vs 9% in patients receiving or not receiving aspirin (p < 0.01 and p = 0.02, respectively, HR 0.4, 95% CI 0.2-0.8 and HR 0.4, 95% CI 0.2-0.9).Additional analyses revealed that the apparent differences in arterial and venous thrombosis observed among type 1/type 1-like CALR-mutated and TN cases, were fully accounted for by aspirin use (p value adjusted for aspirin use = 0.40/0.31).Cytoreductive therapy, on the other hand, did not appear to have a clear beneficial impact on neither arterial (p = 0.08) nor venous thrombosis (p = 0.19).

DISCUSSION
The current study constitutes the largest single-center series of ET patients who are fully annotated for driver mutations and includes mature survival data and detailed analysis of prognostic factors for overall, leukemia-free, myelofibrosis-free, and thrombosis-free survival, with the latter stratified into arterial vs. venous events.Our study provides baseline clinical and laboratory data and confirms previously recognized differences in age and gender distribution as well as hemoglobin, leukocyte, and platelet levels, among specific driver mutation categories; CALR-mutated and TN patients were younger at diagnosis (median age 52 and 50 years, respectively), compared to JAK2 or MPL-mutated cases (median age 71 and 66 years, respectively); JAK2 and TN patients were predominantly female, compared to CALR and MPL-mutated cases [19].Noteworthy laboratory associations included higher hemoglobin and leukocyte count with JAK2 mutation and higher platelet count with CALR mutation (type-2 more than type-1) and TN [19].At the time of diagnosis, approximately 22% of patients displayed history of major arterial (14%) or venous (10%) thrombosis, 8% major hemorrhage, and 29%, microvascular symptoms; incidences of major thrombosis and hemorrhage were lower in CALR-mutated cases [20].
A major strength of the current study was the availability of long-term follow-up data, which enabled accurate estimation of survival and disease transformation rates; median overall survival was 20.6 years, with 10-year/20-year leukemic transformation and fibrotic progression rates of 1.5%/7.6%and 8%/26%, respectively.As previously noted [9], JAK2/CALR/MPL/TN mutational status did not appear to impact overall or leukemia-free survival in our current ET patient cohort while MPL mutation was associated with a significantly higher rate of progression to myelofibrosis, as per previous reports [21,22].Prominent risk factors for survival in the current ET patient cohort included older age, ANC ≥ 8 × 10 9 /L, ALC < 1.7 × 10 9 /L, male gender, hypertension, and arterial thrombosis history.These observations are in line with those previously communicated [10,23].Application of the recently introduced AAA survival model in ET [10], to the current patient cohort resulted in median survival estimates not reached for low (10-/20year survival rate 98%/80%), 21.3 years for intermediate-1 (10-/20year survival rate 93%/60%), and 10.6 years for high/intermediate-2 (10-/20-year survival rate 54%/7%) risk patients (Fig. 1).
Additional observations from the current study are highlighted by (i) the extremely low incidence of leukemic transformation in the absence of abnormal karyotype and ExT (10-/20-year rate of 1.5%/3%), and (ii) the relatively high rate of fibrotic progression in MPL-mutated patients or those with ANC ≥ 8 × 10(9)/L.It is to be recalled that we have previously reported an association between ExT and inferior overall and leukemia-free survival in young (age <40 years) ET patients [24].However, the association between ExT and leukemic progression in the current study appeared to be limited to patients with JAK2 mutation; this is a noteworthy observation, given that ExT is typically associated with CALR (type-2 more than type-1) and TN mutational status.Interestingly, none of our 84 TN patients with ET experienced leukemic progression.These findings require external validation.The prognostic impact of ANC ≥ 8 × 10 9 /L on MFFS, has not been previously described, and was limited to JAK-mutated cases.Furthermore, in line with prior reports, type 1/type 1-like CALR mutated compared to JAK2mutated, and TN cases were noted to have a significantly higher risk of fibrotic progression [22].The current study did not include  information on other mutations that have previously been shown to adversely affect survival, including SF3B1, SRSF2 U2AF1 TP53 mutations [12].The current study also confirms the higher rates of arterial and venous thrombosis displayed by JAK2/MPL-mutated patients with ET, compared with type 1/type 1-like CALR-mutated and TN, and that this effect was not accounted for by other independent risk factors for thrombosis including age, gender, thrombosis history or cardiovascular risk factors [20].Moreover, a non-significantly higher risk of thrombosis was also observed in CALR-mutated patients in comparison with TN cases; which was no longer apparent after accounting for aspirin use.Together, our findings not only confirm differential thrombosis risk according to driver mutation, but also identify TN patients to have a lower risk of thrombosis, akin to those with CALR mutation.Our observations corroborate the salutary effects of aspirin with respect to both arterial and venous thrombosis, and also suggest possible benefits from aspirin prophylaxis in "very low risk" CALR-mutated patients.
From a practical standpoint, the prognostic information discussed in the current report, regarding survival is not necessarily actionable, since drug therapy in ET has not been shown to be disease-modifying.However, identification of prognostic factors for survival and disease progression is useful for purposes of patient counseling and disease monitoring.On the other hand, risk factor assessment for thrombosis is critical for both primary and secondary prevention measures.In this regard, the observations from the current study underscore the therapeutic value of aspirin for the prevention of both arterial and venous thrombosis.Controlled studies are always preferred over retrospective studies for accurate determination of the optimal therapeutic approach to ameliorate the risk associated with thrombosis and disease progression in ET.In the meantime, we hope the information contained in the current document serves as a complimentary resource for patients and physicians and provides context for the design and interpretation of future clinical trials [12,[25][26][27][28][29].

Fig. 1
Fig. 1 Survival and leukemic/myelofibrotic transformation in essential thrombocythemia.a Overall survival in 985 patients with essential thrombocythemia stratified by driver mutation.b Leukemia-free survival in 985 patients with essential thrombocythemia stratified by driver mutation.c Myelofibrosis-free survival in 985 patients with essential thrombocythemia stratified by driver mutation.

Fig. 2
Fig. 2 Triple A risk model in essential thrombocythemia.

Fig. 3
Fig. 3 Blastic transformation and fibrotic progression in essential thrombocythemia.a Blastic transformation risk model in essential thrombocythemia.b Fibrotic progression risk model in essential thrombocythemia.

Fig. 4
Fig. 4 Arterial and venous thrombosis in essential thrombocythemia.a Arterial Thrombosis-free survival in 985 patients with essential thrombocythemia stratified by driver mutation.b Venous thrombosis-free survival in 985 patients with essential thrombocythemia stratified by driver mutation.

Table 2 .
Univariate and multivariable analysis of associations/risk factors for vascular events (arterial/venous thrombosis and hemorrhage) at or after diagnosis among 1000 patients with essential thrombocythemia (ET), fully annotated for driver mutations.*Cardiovascular risk factor includes the presence of diabetes mellitus, hypertension, or smoking.Red shaded boxes represent variables of significance.Gray boxes represent variables that were not computed.

Table 3 .
Univariate and multivariable analysis of risk factors for disease transformation (myelofibrosis and acute leukemia) and overall survival among 1000 patients with essential thrombocythemia (ET), fully annotated for driver mutations.
*Cardiovascular risk factor includes the presence of diabetes mellitus, hypertension, or smoking.Bold value boxes represent variables of significance.N.Gangat et al.

Table 4 .
Univariate and multivariable analysis of risk factors for vascular events (arterial/venous thrombosis and hemorrhage) after diagnosis among 985 patients with essential thrombocythemia (ET), fully annotated for driver mutations with CALR Bold value boxes represent variables of significance.